Identification and intraspecific variability analysis of secreted and membrane-bound proteins from Echinococcus granulosus

ROSENZVIT, M.C.; KAMENETZKY, L.; MUZULIN, P.M.; GUTIERREZ, A.M.; CAMICIA, F.; GUARNERA, E.A.; MCMANUS, D.P.; NAIDICH, A.

Asahikawa, Japón

Simposio; Taeniasis/Cysticercosis and Echinococcosis International Symposium with Focus on Asia and the Pacific and The Third Congress of Federation of Asian Parasitologists focused on Cestode Zoonoses; 2005

Federation of Asian Parasitologists

The species E. granulosus comprises a number of genetic variants or strains that differ in biological features, such as intermediate host specificity, developmental rate and infectivity to humans. It is very important to analyse the existence of associated genetic variation in secreted and membrane bound (S/M) protein coding genes, since this may improve knowledge on strategies of parasite survival in the specific host, with practical consequences in immunodiagnosis and immunoprophylaxis. We used PCR- Single strand conformation polymorphism (SSCP) analysis followed by sequencing to address the genetic variation of the major secreted antigen, the antigen B, and a number of S/M proteins identified by the signal sequence trap technique. Forty two AgB-related genomic sequences were isolated from five E. granulosus strains and clustered in five genes (B1 to 5) and one pseudogene by maximum parsimony analysis. Differential distribution and frequency of AgB variants among strains were found. The level of nucleotide and amino acid variation observed was higher than the reported so far for coding genes of other helmynth parasites. Reverse transcription (RT)-PCR analysis showed that the expression of AgB variants also differed between strains. The signal sequence trap technique was applied in order to identify other genes coding for S/M proteins. Some of the deduced amino acid sequences of the positive clones showed significant similarity with amino acid and Krebs cycle intermediates transporters, presenilins, vacuolar protein sorter proteins, an E. granulosus EST coding for a secreted protein, and some were completely novel. Intra-specific variability studies by PCR SSCP technique confirmed by sequencing, showed that many of the identified S/M protein coding genes were variable between strains. These results suggest that variation in proteins exposed to the host is a more widespread phenomenon in E. granulosus than previously appreciated. We suggest that this variability should be taken into account for the rational design of diagnosis systems of cystic hydatid disease. The diagnostic value of some of the protein variants detected in E. granulosus strains could be determined through inter-laboratory studies as the recently done by the South American Network for Hydatid Serology.